Crystal of reduced glutathione and method for producing same

ABSTRACT

The present invention provides a crystal of reduced glutathione having excellent powder properties and a method for producing the same. The present invention relates to a crystal of reduced glutathione, wherein the average crystal thickness is 10 μm or more.

CROSS-REFERENCE TO RELATED APPLICATIONS

This patent application is a divisional of copending U.S. patentapplication Ser. No. 16/083,483, filed on Sep. 7, 2018, which is theU.S. national phase of International Patent Application No.PCT/JP2017/009655, filed Mar. 10, 2017, which claims the benefit ofJapanese Patent Application No. 2016-053843, filed on Mar. 17, 2016,which are incorporated by reference in their entireties herein.

TECHNICAL FIELD

The present invention relates to a crystal of reduced glutathione havingexcellent powder properties and a method for producing the same.

BACKGROUND ART

Reduced glutathione (γ-L-glutamyl-L-cysteinyl-L-glycine) is a reducingcompound widely existing in organisms and is known to have adetoxification effect in the liver. Therefore, reduced glutathione hasbeen widely used as a product such as a pharmaceutical product, a healthfood, and a cosmetic product, or a raw material or an intermediatethereof. A crystal of reduced glutathione is known to have two types ofpolymorphisms: α crystal and β crystal (Non-Patent Document 1) and, acrystal which is generally utilized as a product is the α crystalbecause of its physical properties.

However, the α-type crystal of reduced glutathione is likely to become aneedle-like or elongated columnar crystal because the crystal growth indirections other than the long-axis (longitudinal) direction is slow.The ratio of the crystal length to the crystal width becomes largebecause of this property, and as a result, there arises a problem thatthe specific volume is increased and the powder flowability isdeteriorated. In order to improve the powder properties, it is necessaryto allow the crystal to grow not only in the longitudinal direction, butalso in the lateral direction and the height direction.

Patent Document 1 describes the improvement of the powder properties ofreduced glutathione and discloses a method in which a crystal of reducedglutathione is added as a seed crystal to an aqueous solution in whichreduced glutathione is dissolved, followed by stirring, therebyprecipitating reduced glutathione in the aqueous solution.

RELATED ART Patent Document

-   Patent Document 1: Japanese Patent No. 5243963

Non-Patent Document

-   Non-Patent Document 1: Yamasaki, K. et al., Analytical Chemistry,    18(7), pp. 874-878 (1969)

DISCLOSURE OF INVENTION Problems to be Solved by the Invention

The method of Patent Document 1 is a method in which supersaturation israpidly destroyed by adding a seed crystal to an aqueous solutioncontaining reduced glutathione at a high concentration, and thereforehas a problem that the growth of the crystal in directions other thanthe longitudinal direction does not catch up with, and the improvementof the powder properties is insufficient.

In view of this, an object of the present invention is to provide acrystal of reduced glutathione having excellent flowability andpulverizability, and a method for producing the same.

Means for Solving the Problems

The present invention relates to the following (1) to (10).

(1) A crystal of reduced glutathione, wherein the average crystalthickness is 10 μm or more.(2) The crystal described in (1), wherein the ratio L/W of the averagecrystal length L to the average crystal width W is 6.0 or less.(3) The crystal described in (1) or (2), wherein the loose specificvolume is 2.5 mL/g or less.(4) The crystal described in any one of (1) to (3), wherein the densespecific volume is 2.0 mL/g or less.(5) The crystal described in any one of (1) to (4), wherein the angle ofrepose is 50° or less.(6) The crystal described in any one of (1) to (5), wherein the angle ofrupture is 45° or less.(7) The crystal described in any one of (1) to (6), wherein the crystalof reduced glutathione is an α crystal.(8) A method for producing a crystal of reduced glutathione, comprisingallowing a crystal of reduced glutathione to exist in an aqueoussolution containing reduced glutathione, and then adding continuously ordividedly an aqueous solution containing reduced glutathione in asupersaturated state to the aqueous solution, thereby precipitatingand/or growing a crystal of reduced glutathione, and thereaftercollecting the crystal of reduced glutathione contained in the aqueoussolution.(9) The production method described in (8), further comprising a step ofpulverizing the collected crystal of reduced glutathione.(10) The production method described in (8) or (9), wherein the crystalof reduced glutathione is an α crystal.

Effects of the Invention

According to the present invention, a crystal of reduced glutathionehaving excellent flowability and pulverizability, and a method forproducing the same are provided.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 illustrates the change in loose specific volume over time whencrystals of reduced glutathione obtained in Example 3 and ComparativeExample were pulverized. The vertical axis represents the loose specificvolume (mL/g), and the horizontal axis represents the pulverization time(min). In FIG. 1, the white lozenges indicate the results of the crystalof Comparative Example, and the black circles indicate the results ofthe crystal of Example 3.

FIG. 2 illustrates a schematic view of a crystal of reduced glutathioneof the present invention.

EMBODIMENTS FOR CARRYING OUT THE INVENTION 1. Crystal of the PresentInvention

In this description, when a crystal is ascertained three-dimensionallyusing a microscope, the longest axis is referred to as “longitudinalaxis”, and an axis perpendicular to the longitudinal axis is referred toas “lateral axis” (FIG. 2). Further, the length in the longitudinal axisdirection of a crystal, the major axis of the cross section of thelateral axis, and the minor axis of the cross section of the lateralaxis are referred to as “crystal length”, “crystal width”, and “crystalthickness”, respectively (see FIG. 2), and the averages thereof arereferred to as “average length”, “average width”, and “averagethickness”, respectively.

Further, the average length, the average width, and the averagethickness are also referred to as “L”, “W”, and “T”, respectively. TheL, W, and T can be measured by the method described in thebelow-mentioned Analysis Example.

The crystal length and the crystal width can be measured, for example,using a light microscope [DIGITAL MICROSCOPE VHX-900 (KEYENCE)] at ameasurement magnification of 300 to 600 times. For example, the crystallength and the crystal width are measured for 200 crystals, and theaverage length L, the average width W, and the ratio L/W of the averagelength L to the average width W are calculated (FIG. 2, left drawing).

The crystal thickness can be measured, for example, using a scanningelectron microscope [JSM-6510 (manufactured by JEOL Ltd.)] at ameasurement magnification of 250 to 1000 times. For example, as for 50crystals, crystals whose longitudinal axis is perpendicular to thescanning electron microscope field of view are selected, and thethickness of each crystal is measured (FIG. 2, right drawing), and theaverage thickness T is measured. A sample is fixed to an aluminum samplestand with a carbon double-sided tape.

The average crystal thickness (T) of the crystal of reduced glutathioneof the present invention is 10 μm or more, preferably 11 μm or more,more preferably 12 μm or more, most preferably 13 μm or more.

A crystal having a large crystal thickness has excellentpulverizability, and therefore, as the crystal of reduced glutathione, acrystal having a large crystal thickness is preferred, however, as theupper limit of the average thickness of the crystal of reducedglutathione of the present invention, generally 40 μm or less,preferably 35 μm or less, more preferably 30 μm or less, most preferably25 μm or less can be exemplified.

The ratio L/W of the average length L to the average width W of thecrystal of reduced glutathione of the present invention is preferably6.0 or less, more preferably 5.5 or less, further more preferably 5.0 orless, most preferably 4.5 or less.

A crystal having a larger L/W is a needle-like or elongated columnarcrystal, and the specific volume is large, and the powder flowability ispoor. Therefore, the L/W of the crystal of reduced glutathione ispreferably small, however, as the lower limit of L/W, generally 1.0 ormore, preferably 1.1 or more, more preferably 1.2 or more, mostpreferably 1.5 or more can be exemplified.

As the crystal of reduced glutathione of the present invention, acrystal of reduced glutathione in which the average crystal thickness(T) is 10 μm or more, and the L/W of the crystal is preferably 6.0 orless can be exemplified.

The loose specific volume of the crystal of reduced glutathione of thepresent invention is preferably 2.5 mL/g or less, more preferably 2.3mL/g or less, further more preferably 2.2 mL/g or less, most preferably2.0 mL/g or less.

A crystal having a small loose specific volume has an excellent fillingproperty, and is easy to handle in various processing steps, and alsoits transport cost is low. Therefore, the crystal of reduced glutathionepreferably has a small loose specific volume, however, as the lowerlimit of the loose specific volume, generally 1.0 mL/g or more,preferably 1.2 mL/g or more can be exemplified.

Here, the “loose specific volume” refers to a value which is obtained bydividing a volume occupied by a powder by a mass when the powder isfilled in a container and the mass of the powder is measured. The loosespecific volume can be measured, for example, using Multi TesterMT-1001T (manufactured by Seishin Enterprise Co., Ltd.) according to theaccompanying manual under the following measurement conditions.

[Measurement Conditions for Loose Specific Volume]

Sieve: 1.4 mm

Spacer: 30 mm

Vibration width: 0.6 to 0.7 mm

Crystal volume: 40 mL

As the crystal of reduced glutathione of the present invention, acrystal of reduced glutathione in which the average crystal thickness(T) is 10 μm or more, and the loose specific volume is preferably 2.5mL/g or less can be exemplified.

The dense specific volume of the crystal of reduced glutathione of thepresent invention is preferably 2.0 mL/g or less, more preferably 1.8mL/g or less, further more preferably 1.6 mL/g or less, most preferably1.5 mL/g or less.

A crystal having a small dense specific volume has an excellent fillingproperty, and also its transport cost is low. Therefore, the crystal ofreduced glutathione preferably has a small dense specific volume,however, as the lower limit of the dense specific volume, generally 0.8mL/g or more, preferably 1.0 mL/g or more can be exemplified.

Here, the “dense specific volume” refers to a value which is obtained bydividing a volume occupied by a powder by a mass when the powder isfilled in a container and the mass of the powder is measured, andthereafter a given impact is applied to the container. The densespecific volume can be measured, for example, using Multi TesterMT-1001T (manufactured by Seishin Enterprise Co., Ltd.) according to theaccompanying manual under the following measurement conditions.

[Measurement Conditions for Dense Specific Volume]

Tapping rate: 1 tap/sec

Number of taps: 200 taps

As the crystal of reduced glutathione of the present invention, acrystal of reduced glutathione in which the average crystal thickness(T) is 10 μm or more, and the dense specific volume is preferably 2.0mL/g or less can be exemplified.

A difference between the loose specific volume and the dense specificvolume of the crystal of reduced glutathione of the present invention ispreferably 1.2 mL/g or less, more preferably 1.0 mL/g or less. As thelower limit of the difference between the loose specific volume and thedense specific volume, generally 0.1 mL/g or more, preferably 0.2 mL/gor more can be exemplified. Here, the “difference between the loosespecific volume and the dense specific volume” refers to a positivevalue when the dense specific volume is subtracted from the loosespecific volume.

As the crystal of reduced glutathione of the present invention, acrystal of reduced glutathione in which the average crystal thickness(T) is 10 μm or more, the L/W of the crystal is preferably 6.0 or less,and the difference between the loose specific volume and the densespecific volume is preferably 1.2 mL/g or less can be exemplified.

The angle of repose of the crystal of reduced glutathione of the presentinvention is preferably 50° or less, more preferably 48° or less,further more preferably 47° or less, most preferably 45° or less.

A crystal having a large angle of repose cannot be completely dischargedfrom the bottom of a hopper unless the angle of tilt of the bottom ofthe hopper is larger than the angle of repose when discharging thecrystal from the hopper, and therefore, the device is limited andhandling becomes complicated. Further, a crystal having a large angle ofrepose has poor flowability. Therefore, the crystal of reducedglutathione preferably has a small angle of repose, however, as thelower limit of the angle of repose, generally 30° or more, preferably35° or more can be exemplified.

Here, the “angle of repose” refers to an angle formed by a horizontalplane and a generating line of a cone formed with a powder when thepowder is allowed to gently fall onto the horizontal plane through akind of a funnel. The angle of repose can be measured, for example,using Multi Tester MT-1001T (manufactured by Seishin Enterprise Co.,Ltd.) according to the accompanying manual under the followingmeasurement conditions.

[Measurement Conditions for Angle of Repose]

An angle-of-repose table is rotated without giving vibration, angles areread at three sites, and the arithmetic mean thereof is determined to bethe angle of repose.

Sieve: 1.4 mm

Vibration width: 0.6 to 0.7 mm

An angle-of-repose table unit (part number: MT-1028) is used.

As the crystal of reduced glutathione of the present invention, acrystal of reduced glutathione in which the average crystal thickness(T) is 10 μm or more, and the angle of repose is preferably 50° or lesscan be exemplified.

The angle of rupture of the crystal of reduced glutathione of thepresent invention is preferably 45° or less, more preferably 43° orless, further more preferably 42° or less, most preferably 40° or less.

A crystal having a large difference between the angle of rupture and theangle of repose has high floodability and is difficult to control, andtherefore, the difference between the angle of rupture and the angle ofrepose is preferably small, however, as the lower limit of the angle ofrupture, generally 30° or more, preferably 35° or more can beexemplified.

Here, the “angle of rupture” refers to an angle formed by a horizontalplane and a generating line of a cone formed when a given impact isindirectly applied to a cone formed with a powder by allowing the powderto gently fall onto the horizontal plane through a kind of a funnel. Theangle of rupture can be measured, for example, using Multi TesterMT-1001T (manufactured by Seishin Enterprise Co., Ltd.) according to theaccompanying manual under the following measurement conditions.

[Measurement Conditions for Angle of Rupture]

An operation in which a weight attached to the bottom of anangle-of-repose table is slowly lifted under a tapping table and allowedto fall repeatedly for 3 times. In the same manner as the measurementmethod for the angle of repose described above, angles are read at threesites, and the arithmetic mean thereof is determined to be the angle ofrupture.

As the crystal of reduced glutathione of the present invention, acrystal of reduced glutathione in which the average crystal thickness(T) is 10 μm or more, and the angle of rupture is preferably 45° or lesscan be exemplified.

The crystal of reduced glutathione of the present invention may be acrystalline powder including polymorphisms such as an α crystal and a 13crystal, however, as the crystal of reduced glutathione, an α crystal ispreferred. As the crystalline powder, a crystalline powder in which theratio of the α crystal to the total reduced glutathione is generally 95%or more, preferably 97% or more, more preferably 98% or more, furthermore preferably 99% or more, particularly preferably 99.5% or more, mostpreferably 99.9% or more can be exemplified.

2. Production Method of the Present Invention

The production method of the present invention is a method comprisingallowing a crystal of reduced glutathione to exist in an aqueoussolution containing reduced glutathione, and then adding continuously ordividedly an aqueous solution containing reduced glutathione in asupersaturated state to the aqueous solution, thereby precipitatingand/or growing a crystal of reduced glutathione, and thereaftercollecting the crystal of reduced glutathione contained in the aqueoussolution.

The solution containing reduced glutathione may be a solution producedby any production method of a fermentation method, an enzymatic method,an extraction method from a natural product, a chemical synthesismethod, and the like. For example, a solution obtained by removinginsoluble substances from a culture containing reduced glutathioneobtained by culturing a microorganism having an ability to produceglutathione (WO 2008/126784), an aqueous solution containing reducedglutathione obtained by an enzymatic method [Appl. Microbiol.Biotechnol., 66, 233 (2004), JP-A-60-105499, etc.], or the like can beexemplified. More preferably, an aqueous solution of reduced glutathioneobtained by the method described in Example 1 of Japanese Patent No.5243963 can be exemplified.

As the method for allowing a crystal of reduced glutathione to exist inan aqueous solution containing reduced glutathione, for example, amethod in which a crystal of reduced glutathione is crystallized in anaqueous solution containing reduced glutathione by concentrating theaqueous solution containing reduced glutathione to a concentration equalto or higher than the saturation solubility can be exemplified. When thecrystal is crystallized by concentration, stirring may be performed.

The method for concentrating the aqueous solution containing reducedglutathione is not particularly limited, and for example, evaporationunder reduced pressure conditions or a method using a reverse osmosismembrane can be exemplified.

As the concentration of reduced glutathione in the aqueous solutioncontaining reduced glutathione when a crystal of reduced glutathione iscrystallized, generally 100 g/L or more, preferably 250 g/L or more,more preferably 400 g/L or more can be exemplified.

Further, as the method for allowing a crystal of reduced glutathione toexist in an r aqueous solution containing reduced glutathione, forexample, a method in which before a crystal of reduced glutathione iscrystallized by concentrating the aqueous solution containing reducedglutathione, a crystal of reduced glutathione is added as a seed crystalso that the concentration in the aqueous solution containing reducedglutathione becomes generally 0.05 to 25 g/L, preferably 0.1 to 10 g/L,thereby crystallizing the crystal can be exemplified. When the crystalis crystallized by adding a seed crystal, stirring may be performed.

As the temperature when the crystal of reduced glutathione iscrystallized, generally 0 to 50° C., preferably 5 to 40° C., morepreferably 10 to 30° C. can be exemplified.

By adding continuously or dividedly an aqueous solution containingreduced glutathione in a supersaturated state to the aqueous solution inwhich the crystal of reduced glutathione is allowed to exist, thecrystal of reduced glutathione can be precipitated and/or grown.

The “precipitating and/or growing the crystal of reduced glutathione”includes 1) newly crystallizing a crystal of reduced glutathione in theaqueous solution containing reduced glutathione, 2) enlarging thecrystallized crystal, and 3) enlarging the crystal of reducedglutathione allowed to exist in the aqueous solution containing reducedglutathione before adding the aqueous solution containing reducedglutathione in a supersaturated state, by adding the aqueous solutioncontaining reduced glutathione in a supersaturated state.

The aqueous solution containing reduced glutathione in a supersaturatedstate can be prepared by the same method as described above.

The temperature at which the aqueous solution containing reducedglutathione in a supersaturated state is added is not particularlylimited as long as reduced glutathione is not precipitated, however,generally 0 to 50° C., preferably 5 to 40° C., more preferably 10 to 30°C. can be exemplified.

The addition of the aqueous solution containing reduced glutathione in asupersaturated state may be performed continuously at a given rate, ormay be performed by dividing the total liquid amount into portions.

When the aqueous solution containing reduced glutathione in asupersaturated state is added in divided portions, the aqueous solutioncontaining reduced glutathione in a supersaturated state can be dividedinto portions and added an arbitrary number of times, and the respectivedivided portions can be added at arbitrary intervals. Further, when thereduced glutathione-containing aqueous solution in a supersaturatedstate is not added, only stirring may be continued.

As the time required for adding the aqueous solution containing reducedglutathione in a supersaturated state, generally 4 to 50 hours,preferably 7 to 40 hours, more preferably 10 to 30 hours can beexemplified.

As the amount of the aqueous solution containing reduced glutathione ina supersaturated state to be added, generally 2 to 200 times equivalent,preferably 5 to 100 times equivalent, more preferably 8 to 50 timesequivalent with respect to the aqueous solution containing reducedglutathione in which the crystal of reduced glutathione is crystallizedcan be exemplified.

By cooling after adding the reduced aqueous solution containing reducedglutathione, the precipitation and/or growth of the crystal ofglutathione can be accelerated. As the cooling temperature, generally40° C. or lower, preferably 30° C. or lower, more preferably 20° C. orlower can be exemplified.

After adding the aqueous solution containing reduced glutathione in asupersaturated state, by adding or dropping a solvent selected from thegroup consisting of alcohols and ketones as needed, the precipitationand/or growth of the crystal of reduced glutathione can be accelerated.

It is also possible to use a solution obtained by mixing an alcohol or aketone with water at an arbitrary ratio in place of the solvent selectedfrom the group consisting of alcohols and ketones.

As the alcohols, preferably C1 to C6 alcohols, more preferably C1 to C3alcohols, further more preferably alcohols selected from the groupconsisting of methanol, ethanol, n-propanol, and isopropyl alcohol, mostpreferably alcohols selected from the group consisting of methanol andethanol can be exemplified.

As the ketones, preferably ketones selected from acetone, methyl ethylketone, and diethyl ketone, more preferably acetone can be exemplified.

The temperature when the alcohols and the ketones are added or droppedmay be any temperature as long as it is a temperature at which reducedglutathione is not decomposed, however, in order to improve thecrystallization ratio of the crystal of reduced glutathione bydecreasing the solubility, generally 40° C. or lower, preferably 30° C.or lower, more preferably 25° C. or lower, most preferably 20° C. orlower can be exemplified. As the lower limit of the temperature,generally 0° C. or higher, preferably 5° C. or higher can beexemplified.

As the amount of the alcohols and the ketones to be added or dropped,generally 0.1 to 3 times, preferably 0.2 to 2 times the amount of theaqueous solution can be exemplified.

After the crystal of reduced glutathione is precipitated and/or grown asdescribed above, the aqueous solution containing the grown crystal isfurther stirred or left as such at generally 0 to 40° C., preferably 5to 30° C., more preferably 5 to 20° C. for generally 1 to 48 hours,preferably 1 to 24 hours, most preferably 1 to 12 hours, whereby thecrystal can be aged.

The “aging” refers to further growing the crystal by stopping the stepof precipitating the crystal of reduced glutathione.

The aging of the crystal is performed for growing the crystal as themain purpose, however, simultaneously with the growth of the crystal,precipitation of a new crystal may occur.

After aging the crystal, the step of precipitating and/or growing thecrystal of reduced glutathione may be restarted.

As the method for collecting the crystal of reduced glutathione obtainedby precipitating and/or growing the crystal or the crystal from theaqueous solution containing the crystal of reduced glutathione obtainedby further aging in the above, for example, collection by filtration,pressure filtration, suction filtration, centrifugation, and the likecan be exemplified. Further, in order to reduce the adhesion of themother liquor to the crystal so as to improve the quality of thecrystal, after collecting the crystal, the crystal may be washed asappropriate.

A solution to be used for washing is not particularly limited, however,water, methanol, ethanol, acetone, n-propanol, isopropyl alcohol, andone type of solution selected therefrom, or a solution obtained bymixing a plurality of types selected therefrom at an arbitrary ratio canbe used.

A wet crystal obtained in the above may be dried. The drying conditionmay be any condition as long as it is a method capable of maintainingthe form of the crystal of reduced glutathione, and for example, reducedpressure drying, vacuum drying, fluidized bed drying, ventilationdrying, and the like can be exemplified.

The drying temperature may be any temperature as long as adhesive wateror a solution can be removed, and reduced glutathione is not decomposed,however, generally, 70° C. or lower, preferably 60° C. or lower, morepreferably 50° C. or lower can be exemplified.

The crystal of reduced glutathione of the present invention can beproduced by further pulverizing the crystal of reduced glutathione ofthe present invention obtained by the above-mentioned method. Thepulverization of the crystal can be performed, for example, usingOsterizer Oster Vintage Blender 16-speed Dual Range (Osterizer OSTER)under the following conditions.

[Conditions for Pulverization of Crystal]

Rotation speed: 33700 to 33900 r/min

Diameter: 50 mm

Sample feeding amount: 20 g/feed

EXAMPLES

Hereinafter, Examples will be shown, however, the present invention isnot limited to the following Examples.

Example 1 Production of Crystal of the Present Invention (1)

An aqueous solution containing reduced glutathione at a concentration of125 g/L was obtained according to the method described in Example 1 ofJapanese Patent No. 5243963. The aqueous solution was concentrated to530 g/L by heating under reduced pressure. To 180 mL of the concentratedsolution at 25° C., 0.02 g of an α-type crystal of reduced glutathione(manufactured by Kojin Co., Ltd.) was added as a seed crystal, followedby stirring, whereby the α-type crystal of reduced glutathione wascrystallized.

To the aqueous solution, 3850 mL of an aqueous solution containingreduced glutathione concentrated to 520 to 530 g/L was added at 25° C.over 17 hours, whereby the crystal was precipitated and/or grown. Theobtained aqueous solution containing the α-type crystal of reducedglutathione was cooled to 10° C., and 0.3 times equivalent of ethanolwas added thereto, and thereafter, the α-type crystal of reducedglutathione obtained by removing the aqueous solution layer throughcentrifugation was washed with 30 v/v % ethanol, and then dried byventilation at 40° C., whereby the α-type crystal of reduced glutathionewas obtained.

Example 2 Production of Crystal of the Present Invention (2)

An aqueous solution containing reduced glutathione obtained in the samemanner as in Example 1 was concentrated to 436 g/L by heating underreduced pressure. To 330 mL of the concentrated solution at 25° C., 0.02g of an α-type crystal of reduced glutathione (manufactured by KojinCo., Ltd.) was added as a seed crystal, followed by stirring, wherebythe α-type crystal of reduced glutathione was crystallized. To theaqueous solution, 2920 mL of a reduced glutathione aqueous solutionconcentrated to 420 to 430 g/L was added at 25° C. over 11 hours,whereby the crystal was precipitated and/or grown.

The obtained aqueous solution containing the α-type crystal of reducedglutathione was cooled to 10° C., and 0.3 times equivalent of ethanolwas added thereto, and thereafter, the α-type crystal of reducedglutathione obtained by removing the aqueous solution layer throughcentrifugation was washed with 30 v/v % ethanol, and then dried byventilation at 40° C., whereby the α-type crystal of reduced glutathionewas obtained.

Example 3 Production of Crystal of the Present Invention (3)

An aqueous solution containing reduced glutathione at a concentration of134 g/L was prepared in the same manner as in Example 1, and thereafterconcentrated to 540 g/L by heating under reduced pressure. To 190 mL ofthe concentrated solution at 25° C., 0.02 g of an α-type crystal ofglutathione (manufactured by Kojin Co., Ltd.) was added as a seedcrystal, followed by stirring, whereby the α-type crystal of reducedglutathione was crystallized.

To the aqueous solution, 3920 mL of a reduced glutathione aqueoussolution concentrated to 540 to 550 g/L was added at 25° C. over 15hours, whereby the crystal was precipitated and/or grown. The obtainedaqueous solution containing the α-type crystal of reduced glutathionewas cooled to 10° C., and 0.3 times equivalent of ethanol was addedthereto, and thereafter, the α-type crystal of reduced glutathioneobtained by removing the aqueous solution layer through centrifugationwas washed with 30 v/v % ethanol, and then dried under reduced pressureat room temperature using a box can, whereby the α-type crystal ofreduced glutathione was obtained.

COMPARATIVE EXAMPLE

An aqueous solution containing reduced glutathione at a concentration of179 g/L was obtained according to the method described in Example 1 ofJapanese Patent No. 5243963. The aqueous solution was concentrated to546 g/L by heating under reduced pressure. To 950 mL of the concentratedsolution at 25° C., 0.05 g of an α-type crystal of reduced glutathione(manufactured by Kojin Co., Ltd.) was added as a seed crystal. After theseed crystal was added, stirring was performed at 25° C. for 10 hours,whereby the α-type crystal of reduced glutathione was crystallized.

To the obtained aqueous solution containing the α-type crystal ofreduced glutathione was added 0.3 times equivalent of ethanol, themixture was cooled to 10° C., and thereafter, the α-type crystal ofreduced glutathione obtained by removing the aqueous solution layerthrough centrifugation was washed with 60 v/v % ethanol, and then driedunder reduced pressure at 40° C., whereby the α-type crystal of reducedglutathione was obtained.

Example 4 Measurement of Powder Properties

With respect to the α-type crystals of reduced glutathione obtained inExamples 1 to 3, the α-type crystal of reduced glutathione obtained inComparative Example, and commercially available α-type crystals ofreduced glutathione (commercially available products A and B), theaverage length L, the average width W, the ratio L/W of the averagelength L to the average width W, the loose specific volume, and thedense specific volume of the crystal were measured. The results areshown in Table 1.

TABLE 1 Loose Dense Difference specific specific in specific L W L/Wvolume volume volume Sample [μm] [μm] [—] [mL/g] [mL/g] [mL/g] Commer-24 to 33 3 to 4 7 to 9 4.69 2.49 2.20 cially available product A Commer-46 to 55 4 to 5 11 to 15 5.50 2.88 2.61 cially available product BCompar- 77 to 94 10 to 11  8 to 11 3.42 1.95 1.48 ative Example Example1 75 to 80 27 to 29 2.7 to 3.2 1.88 1.26 0.62 Example 2 49 to 57 24 to27 2.0 to 2.4 2.00 1.29 0.71 Example 3 67 to 95 23 to 26 2.9 to 4.1 2.001.29 0.71

As shown in Table 1, it was found that in the case of the crystals ofthe present invention obtained in Examples 1 to 3, the L/W, the loosespecific volume, the dense specific volume, and the difference betweenthe loose specific volume and the dense specific volume are all smaller,and the flowability is higher, and the powder properties are superior ascompared with the crystal of glutathione obtained in Comparative Exampleand the commercially available products A and B.

In addition, the average crystal thickness T of each of the α-typecrystals of reduced glutathione obtained in Examples 1 to 3, the α-typecrystal of reduced glutathione obtained in Comparative Example, and thecommercially available product A was measured. The results are shown inTable 2.

TABLE 2 Average crystal thickness T Sample [μm] Commercially availableproduct A 1.9 Comparative Example 5.5 Example 1 17.0 Example 2 16.7Example 3 16.7

As shown in Table 2, it was found that in the case of the α-typecrystals of reduced glutathione obtained in Examples 1 to 3, the averagecrystal thickness T is larger, and the pulverizability is superior ascompared with the α-type crystal of reduced glutathione obtained inComparative Example and the commercially available product A.

Example 5 Production of Crystal of the Present Invention (4)

The α-type crystals of reduced glutathione obtained in Example 3 andComparative Example were pulverized, and the change in loose specificvolume over time was measured.

As a result, it was found that the α-type crystal of reduced glutathioneobtained in Example 3 is pulverized more promptly than the α-typecrystal of reduced glutathione obtained in Comparative Example and thespecific volume thereof converges to a small value (FIG. 1).

Further, the loose specific volume of the α-type crystal of reducedglutathione obtained in Example 3 was smaller than that of the α-typecrystal of reduced glutathione obtained in Comparative Example in allpulverization times.

Further, with respect to the α-type crystal of reduced glutathioneobtained in Example 3 and the α-type crystal of reduced glutathioneobtained in Comparative Example, the angle of repose and the angle ofrupture were measured. The results are shown in Table 3.

TABLE 3 Angle of repose Angle of rupture Sample [°] [°] ComparativeBefore 51.4 46.8 Example pulverization After pulverization 51.8 41.5Example 3 Before 42.4 39.4 pulverization After pulverization 37.4 31.1

As shown in Table 3, it was found that in the case of the α-type crystalof reduced glutathione obtained in Example 3, the angle of repose andthe angle of rupture are smaller than those of the α-type crystal ofreduced glutathione obtained in Comparative Example both before andafter pulverization, and therefore, the flowability is high and thepowder properties are excellent.

From the above-mentioned results, it was found that the crystals of thepresent invention have excellent pulverizability and powder propertiesas compared with the currently available crystals of reducedglutathione.

While the present invention has been described in detail with referenceto specific embodiments, it is apparent to those skilled in the art thatvarious changes and modifications can be made without departing from thespirit and scope of the present invention. The present application isbased on Japanese Patent Application (Japanese Patent Application No.2016-53843) filed on Mar. 17, 2016 and the entire contents of which areincorporated herein by reference. Further, all references cited hereinare incorporated herein by reference in their entirety.

INDUSTRIAL APPLICABILITY

According to the present invention, a crystal of reduced glutathionehaving excellent powder properties and a method for producing the sameare provided.

1. A method for producing a crystal of reduced glutathione, comprisingallowing a crystal of reduced glutathione to exist in an aqueoussolution containing reduced glutathione, and then adding continuously ordividedly an aqueous solution containing reduced glutathione in asupersaturated state to the aqueous solution, thereby precipitatingand/or growing a crystal of reduced glutathione, and thereaftercollecting the crystal of reduced glutathione contained in the aqueoussolution.
 2. The production method according to claim 1, furthercomprising a step of pulverizing the collected crystal of reducedglutathione.
 3. The production method according to claim 2, wherein thecrystal of reduced glutathione is an α crystal.
 4. The production methodaccording to claim 1 wherein the crystal of reduced glutathione is an αcrystal.